Over the past year, studies have continued using murine retroviral vectors that either express the NeoR gene or a mutated, non-expressing NeoR gene. In addition, studies have been initiated using human lentiviral and murine retroviral vectors constructed to express enhanced green fluorescent protein (EGFP). The EGFP expressing vectors have been used to evaluate and optimize transduction protocols targeting immunoselected CD34+ cells. We have demonstrated that a commercially available RGD-containing fibronectin fragment, RetroNectin, improves the efficiency of transduction of CD34+ cells for VSV-G pseudotyped human lentiviral vectors, as well as for murine retroviral vectors that use either GaLV or amphotropic envelope protein. In addition, transplantation experiments using cytokine cocktails containing SCF and flt-3 ligand, in combination with either autologous stroma or RetroNectin, also improves transduction of CD34+ cells that contribute to the hematopoietic reconstitution of transplanted non-human primates. Comparative studies have also been performed using expressing and non-expressing NeoR gene, assessing the longevity of genetically marked peripheral blood (PB) lymphocytes and cells derived from cytokine mobilized immunoselected PB CD34+ cells. A variety of transduction protocols have been evaluated. There is a difference in lymphocyte longevity between mature lymphocytes and the lymphocytic progeny of immunoselected CD34+ cells transduced with a NeoR expressing vector: genetically marked lymphocytes derived from immunoselected CD34+ appear to have a longer lifespan in the circulation than do mature lymphocytes transduced with the identical vector. Studies also have now been initiated evaluating the rhesus macaque as an animal model for the study of Factor IX deficiency, or Hemophilia B. The rhesus macaque Factor IX gene was cloned and sequenced, permitting comparative studies between human and non-human primate Factor IX: the sequence similarity between rhesus and human Factor IX is over 95%. Administration of human Factor IX to non-human primates elicited little to no antibody response, and gene therapy studies are in progress evaluating intravenous administration of an adenoviral vector constructed to express Factor IX. Other studies that continue outside of the gene therapy program include the determination of the best adjuvant for a B19 capsid vaccine, attempts to isolate potential viral agents that may be associated with hepatitis in aplastic anemia patients in tamarins, and exploring the potential use of the rhesus macaque as an animal model for testing biological or chemical agents that may prove beneficial in hemoglobinopathies for stimulation of F-cell expansion and HbF levels.